The signal transducer and activator of transcription 5A (STAT5A) is a transcription factor that mediates the function of various hormones and cytokines, including growth hormone (GH). Mutations in the STAT5A gene, such as the substitution of a G by a C in exon 8, have been associated with differences in production and reproductive performance of dairy cows. The aim of this study was to evaluate the effect of STAT5A BstEII polymorphism on the days from calving to first ovulation and calving-conception interval (CCI). For identification of the polymorphisms DNA was extracted from blood and a fragment of the STAT5A gene was amplified by PCR. The presence of the G and C alleles was determined after digestion of the PCR products with the BstEII enzyme and gel electrophoresis. For this study 73 Holstein cows were followed from 21 d prepartum to 210 d in milk (DIM). At 55 DIM the cows were submitted to an OvSynch-TAI protocol, which was repeated in cows diagnosed as not pregnant. From calving, milk production was recorded, milk samples for progesterone measurement were collected twice a week to determine ovulation day until 60 DIM, and the CCI was evaluated until 210 DIM. Serum samples for insulin-like growth factor I (IGF-I) measurement were collected at −21, 0, 7, 21 and 60 DIM. Data were analyzed using the GLM procedure of SAS. In total, 19 cows (26%) were of the CC genotype, 32 cows (43.8%) of the GC genotype and 22 cows (30.2%) of the GG genotype. The calving to ovulation interval was not different between genotypes (P > 0.05): 28.4 ± 3.1, 29.5 ± 2.5 and 29.4 ± 2.9 DIM for the CC, CG and GG genotypes. The CCI was 101.3 ± 9.4, 102.7 ± 7.6 and 93.9 ± 9.4 DIM for the CC, CG and GG genotypes (P > 0.05). Milk production was similar between the 3 genotypes (P > 0.05). Serum IGF-I was also not different between genotypes, being 66.3 ± 7.2, 61.8 ± 5.8, 65.6 ± 6.7 ng/mL for CC, CG and GG genotypes (P > 0.05). Therefore, the STAT5A BstEII polymorphism did not affect the calving to ovulation interval, CCI, milk production or IGF-I concentrations in Holstein dairy cows. It should be noticed that this study used a small number of cows and larger studies are necessary to confirm current results.